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Background: Antibody-mediated rejection (AMR) is one of the most important challenges in the context of renal transplantation, because the binding of de novo donor-specific antibodies (dnDSA) to the kidney graft triggers the activation of the complement, which in turn leads to loss of transplant. In this context, the objective of this study was to evaluate the association between complement-fixing dnDSA antibodies and graft loss as well as the possible association between non-complement-fixing antibodies and transplanted organ survival in kidney transplant recipients. Methods: Our study included a cohort of 245 transplant patients over a 5-year period at Virgen de las Nieves University Hospital (HUVN) in Granada, Spain. Results: dnDSA was observed in 26 patients. Of these patients, 17 had non-complement-fixing dnDSA and 9 had complement-fixing dnDSA. Conclusions: Our study demonstrated a significant association between the frequency of rejection and renal graft loss and the presence of C1q-binding dnDSA. Our results show the importance of the individualization of dnDSA, classifying them according to their ability to activate the complement, and suggest that the detection of complement-binding capacity by dnDSA could be used as a prognostic marker to predict AMR outcome and graft survival in kidney transplant patients who develop dnDSA.

Chronic lymphocytic leukemia (CLL) is the most common leukemia among adults worldwide. Although genome-wide association studies (GWAS) have uncovered the germline genetic component underlying CLL susceptibility, the potential use of GWAS-identified risk variants to predict disease progression and patient survival remains unexplored. Here, we evaluated whether 41 GWAS-identified risk variants for CLL could influence overall survival (OS) and disease progression, defined as time to first treatment (TTFT) in a cohort of 1039 CLL cases ascertained through the CRuCIAL consortium. Although this is the largest study assessing the effect of GWAS-identified susceptibility variants for CLL on OS, we only found a weak association of ten single nucleotide polymorphisms (SNPs) with OS (p < 0.05) that did not remain significant after correction for multiple testing. In line with these results, polygenic risk scores (PRSs) built with these SNPs in the CRuCIAL cohort showed a modest association with OS and a low capacity to predict patient survival, with an area under the receiver operating characteristic curve (AUROC) of 0.57. Similarly, seven SNPs were associated with TTFT (p < 0.05); however, these did not reach the multiple testing significance threshold, and the meta-analysis with previous published data did not confirm any of the associations. As expected, PRSs built with these SNPs showed reduced accuracy in prediction of disease progression (AUROC = 0.62). These results suggest that susceptibility variants for CLL do not impact overall survival and disease progression in CLL patients, Horizon 2020 856620, Instituto de Salud Carlos III Spanish Government, Marie Curie Actions PI17/02256 PI20/01845, Consejeria de Transformacion Economica, Industria, Conocimiento y Universidades y FEDER PY20/01282, United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Cancer Institute (NCI) P50 CA97274 R01 CA92153

Background. The influence of HLA class I and II loci on the susceptibility to melanoma remains an area of intense debate. This study aimed to examine whether the HLA system was related to melanoma susceptibility and prognosis in a southern Spanish population. Methods. In this study, HLA class I and class II genotyping were performed using polymerase chain reaction sequence-specific oligonucleotides (PCR-SSO) in 237 Spanish melanoma patients and 636 ethnically matched controls. Data were analyzed according to the clinical characteristics of the defined subgroups. Results. Compared to the control group, DRB1 ∗ 16:01 (4% vs. 1.3%, p = 0.001 , Pc = 0.035, OR = 3.28) and DQB1 ∗ 05:02 (4.9% vs. 2%, p = 0.001 , Pc = 0.017, OR = 2.54) were positivity associated with the susceptibility to melanoma. Both DRB1 ∗ 16:01 (5.4% vs. 1.3%, p = 0.001 , Pc = 0.035, OR = 4.46) and DQB1 ∗ 05:02 (6.5% vs. 2%, p = 0.001 , Pc = 0.017, OR = 3.44) also showed a positive correlation with Breslow thickness >1.5 mm, most notably at an early age of diagnosis (≤58 years), DRB1 ∗ 16:01 (4.2% vs. 1.3%, p = 0.001 , Pc = 0.035, OR = 3.41) and DQB1 ∗ 05:02 (5.4% vs. 2%, p = 0.002 , Pc = 0.034, OR = 2.86). Conclusion. These findings established HLA-DRB1 ∗ 16:01 and HLA-DQB1 ∗ 05:02 loci as melanoma risk factors in the southern Spanish population.

The SARS-CoV-2 disease presents different phenotypes of severity. Comorbidities, age, and being overweight are well established risk factors for severe disease. However, innate immunity plays a key role in the early control of viral infections and may condition the gravity of COVID-19. Natural Killer (NK) cells are part of innate immunity and are important in the control of virus infection by killing infected cells and participating in the development of adaptive immunity. Therefore, we studied the short tandem repeat (STR) transmembrane polymorphisms of the major histocompatibility complex class I chain-related A (MICA), an NKG2D ligand that induces activation of NK cells, among other cells. We compared the alleles and genotypes of MICA in COVID-19 patients versus healthy controls and analyzed their relation to disease severity. Our results indicate that the MICA*A9 allele is related to infection as well as to symptomatic disease but not to severe disease. The MICA*A9 allele may be a risk factor for SARS-CoV-2 infection and symptomatic disease., Instituto de Salud Carlos III - FEDER funds (European Union) PI 16/00752 B-CTS-410-UGR-20, Junta de Andalucia CTS-143 C-0013-2018

Acknowledgments: We thank Per Anderson for the English revision. This study is part of the doctoral thesis of Irene Díaz Alberola, within the program of Biomedicine, conducted at the University of Granada, Spain., Institutional Review Board Statement: The study was conducted in accordance with the Declaration of Helsinki, and approved by Portal de Ética de la Investigación Biomédica. Junta de Andalucía (Code: 0297-N-21)., Background and Aims: Common variable immunodeficiency (CVID) comprises a group of diseases with heterogeneous clinical and immunological manifestations. Several mutations have been identified in genes encoding proteins essential for immune function. Our aim was to phenotypically and genotypically characterize a patient diagnosed with CVID and study his response to the SARSCoV- 2 vaccine. Methods: We performed a next-generation sequencing analysis, a CMIA, and an ELISA to analyze the humoral and cellular response to the SARS-CoV-2 vaccine, respectively. We also employed flow cytometry and immunoturbidimetry to assess the patient’s global immune status. Results: We found a low humoral but positive cellular response to the SARS-CoV-2 vaccine. NGS screening revealed a transition from guanine to adenine at position c.485 of the IKZF1 gene in heterozygosity, giving rise to the R162Q variant, which was not present in his parents. Conclusions: The R162Q variant of the IKZF1 gene has been associated with CVID type 13, but always with an autosomal dominant inheritance with high penetrance. Therefore, we present for the first time a case of CVID associated with a de novo heterozygous R162Q variant in the IKZF1 gene in a patient with a low humoral immune response to the complete COVID-19 vaccination program., Partially financed by Palex Medical S.A.

Common variable immunodeficiency (CVID) comprises a group of diseases with heterogeneous clinical and immunological manifestations. Several mutations have been identified in genes encoding proteins essential for immune function. Our aim was to phenotypically and genotypically characterize a patient diagnosed with CVID and study his response to the SARS-CoV-2 vaccine.We performed a next-generation sequencing analysis, a CMIA, and an ELISA to analyze the humoral and cellular response to the SARS-CoV-2 vaccine, respectively. We also employed flow cytometry and immunoturbidimetry to assess the patient's global immune status.We found a low humoral but positive cellular response to the SARS-CoV-2 vaccine. NGS screening revealed a transition from guanine to adenine at position c.485 of theThe R162Q variant of the

This study was supported by grants from the FIBAO foundation (Granada, Spain) and from the Instituto de Salud Carlos III (PI12/02688, PI17/02256 and PI20/01845; Madrid, Spain)., In this study, we have evaluated whether 57 genome-wide association studies (GWAS)-identified common variants for type 2 diabetes (T2D) influence the risk of developing prostate cancer (PCa) in a population of 304 Caucasian PCa patients and 686 controls. The association of selected single nucleotide polymorphisms (SNPs) with the risk of PCa was validated through meta-analysis of our data with those from the UKBiobank and FinnGen cohorts, but also previously published genetic studies. We also evaluated whether T2D SNPs associated with PCa risk could influence host immune responses by analysing their correlation with absolute numbers of 91 blood-derived cell populations and circulating levels of 103 immunological proteins and 7 steroid hormones. We also investigated the correlation of the most interesting SNPs with cytokine levels after in vitro stimulation of whole blood, peripheral mononuclear cells (PBMCs), and monocyte-derived macrophages with LPS, PHA, Pam3Cys, and Staphylococcus Aureus. The meta-analysis of our data with those from six large cohorts confirmed that each copy of the FTOrs9939609A, HNF1B(rs7501939T), HNF1B(rs75721T), HNF1B(rs4430796G), and JAZF1(rs10486567A) alleles significantly decreased risk of developing PCa (p = 3.70 x 10(-5), p = 9.39 x 10(-54), p = 5.04 x 10(-54), p = 1.19 x 10(-71), and p = 1.66 x 10(-18), respectively). Although it was not statistically significant after correction for multiple testing, we also found that the NOTCH2(rs10923931T) and RBMS1(rs7593730) SNPs associated with the risk of developing PCa (p = 8.49 x 10(-4) and 0.004). Interestingly, we found that the protective effect attributed to the HFN1B locus could be mediated by the SULT1A1 protein (p = 0.00030), an arylsulfotransferase that catalyzes the sulfate conjugation of many hormones, neurotransmitters, drugs, and xenobiotic compounds. In addition to these results, eQTL analysis revealed that the HNF1B(rs7501939), HNF1B(rs757210), HNF1B(rs4430796), NOTCH2(rs10923931), and RBMS1(rs7593730) SNPs influence the risk of PCa through the modulation of mRNA levels of their respective genes in whole blood and/or liver. These results confirm that functional TD2-related variants influence the risk of developing PCa, but also highlight the need of additional experiments to validate our functional results in a tumoral tissue context., FIBAO foundation (Granada, Spain), Instituto de Salud Carlos III PI12/02688 PI17/02256 PI20/01845

This work was supported by "Investigacion y Desarrollo (I + D) del Sistema Andaluz de Salud (SAS)" and Instituto de Salud Carlos III (Proyecto FIS PI21/01708)., The new vaccines against SARS-CoV-2 have raised a lot of expectations about their ability to induce immunity and the duration of this. This is the case of mRNA vaccines such as Moderna’s mRNA-1273. Therefore, it is necessary to study the humoral and cellular immunity generated by these vaccines. Our objectives are determining what is the normal response of antibody production, and what is the level of protective antibodies and monitoring patients in case of subsequent infection with COVID-19. We present the first results of a longitudinal study of the humoral response in 601 health workers vaccinated with Moderna. The results show a humoral immunity at 90 days after the second dose of 100%, with a strong decrease between the levels of circulating anti-S IgG antibodies between days 30 and 90 postvaccination. Observing a steeper decline in those who had higher titles at the beginning. In addition, we present a cellular response of 86% at three months after the second dose, which is related to low humoral response., Investigacion y Desarrollo (I + D) del Sistema Andaluz de Salud (SAS), Instituto de Salud Carlos III European Commission FIS PI21/01708

Common variable immunodeficiency (CVID) is an antibody immunodeficiency with a wide variety of clinical and immunological manifestations, and whose genetic cause is found in about 25% of diagnosed cases. Giardia lamblia is one of the main causes of gastrointestinal infections in CVID. 5-Nitroimidazoles are the most used first-line treatment, but nitroimidazole-refractory giardiasis is increasing. Nevertheless, only a few cases of refractory giardiasis in CVID have been reported. This study aimed to determine the incidence of Giardia infection in our CVID cohort, shows our management experience and describes patients’ phenotypic features. Clinical data collection, immunological, immunogenetics and microbiology assays were performed, and previous cases of giardiasis in CVID were reviewed. The incidence of symptomatic giardiasis was 12.9%. The main immunological features were undetectable or decreased IgA levels and reduced switched memory B cells. A probable PTEN pathogenic variant was detected in one. Three patients responded to metronidazole but suffered reinfections, and one was a refractory giardiasis eradicated with innovative quinacrine plus paromomycin combination. This work could contribute to the decision-making and therapeutic management of future patients with CVID and giardiasis, highlighting the importance of the early detection and treatment of infections in patients with CVID to ensure a good quality of life.

Autophagy is a highly conserved metabolic pathway via which unwanted intracellular materials, such as unfolded proteins or damaged organelles, are digested. It is activated in response to conditions of oxidative stress or starvation, and is essential for the maintenance of cellular homeostasis and other vital functions, such as differentiation, cell death, and the cell cycle. Therefore, autophagy plays an important role in the initiation and progression of tumors, including hematological malignancies, where damaged autophagy during hematopoiesis can cause malignant transformation and increase cell proliferation. Over the last decade, the importance of autophagy in response to standard pharmacological treatment of hematological tumors has been observed, revealing completely opposite roles depending on the tumor type and stage. Thus, autophagy can promote tumor survival by attenuating the cellular damage caused by drugs and/or stabilizing oncogenic proteins, but can also have an antitumoral effect due to autophagic cell death. Therefore, autophagy-based strategies must depend on the context to create specific and safe combination therapies that could contribute to improved clinical outcomes. In this review, we describe the process of autophagy and its role on hematopoiesis, and we highlight recent research investigating its role as a potential therapeutic target in hematological malignancies. The findings suggest that genetic variants within autophagy-related genes modulate the risk of developing hemopathies, as well as patient survival.

Background: Human leukocyte antigen (HLA) plays an important role in immune responses to infections, especially in the development of acquired immunity. Given the high degree of polymorphisms that HLA molecules present, some will be more or less effective in controlling SARS-CoV-2 infection. We wanted to analyze whether certain polymorphisms may be involved in the protection or susceptibility to COVID-19. Methods: We studied the polymorphisms in HLA class I (HLA-A, -B and -C) and II (HLA-DRB1 and HLA-DQB1) molecules in 450 patients who required hospitalization for COVID-19, creating one of the largest HLA-typed patient cohort to date. Results: Our results show that there is no relationship between HLA polymorphisms or haplotypes and susceptibility or protection to COVID-19. Conclusion: Our results may contribute to resolve the contradictory data on the role of HLA polymorphisms in COVID-19 infection., Instituto de Salud Carlos III - FEDER funds (European Union) PI 16/00752, Junta de Andalucia CTS-143 C-0013-2018, RAFER S.L.

Here, we assessed whether 36 single nucleotide polymorphisms (SNPs) within the TNFSF4 and MAPKAPK2 loci influence the risk of developing invasive aspergillosis (IA). We conducted a twostage case control study including 911 high-risk patients diagnosed with hematological malignancies that were ascertained through the aspBIOmics consortium. The meta-analysis of the discovery and replication populations revealed that carriers of the TNFSF4rs7526628T/T genotype had a significantly increased risk of developing IA (p = 0.00022). We also found that carriers of the TNFSF4rs7526628T allele showed decreased serum levels of TNFSF14 protein (p = 0.0027), and that their macrophages had a decreased fungicidal activity (p = 0.048). In addition, we observed that each copy of the MAPKAPK2rs12137965G allele increased the risk of IA by 60% (p = 0.0017), whereas each copy of the MAPKAPK2rs17013271T allele was estimated to decrease the risk of developing the disease (p = 0.0029). Mechanistically, we found that carriers of the risk MAPKAPK2rs12137965G allele showed increased numbers of CD38+IgM-IgD- plasmablasts in blood (p = 0.00086), whereas those harboring two copies of the allele had decreased serum concentrations of thymic stromal lymphopoietin (p = 0.00097). Finally, we also found that carriers of the protective MAPKAPK2rs17013271T allele had decreased numbers of CD27-IgM-IgD- B cells (p = 0.00087) and significantly lower numbers of CD14+ and CD14+CD16- cells (p = 0.00018 and 0.00023). Altogether, these results suggest a role of the TNFSF4 and MAPKAPK2 genes in determining IA risk., Instituto de Salud Carlos III PI17/02276, ERA-NET PathoGenoMics PIM2010EPA-00756 0315900A, Collaborative Research Center/Transregio 124 FungiNet, Portuguese Foundation for Science and Technology PTDC/SAU-SER/29635/2017 PTDC/MED-GEN/28778/2017 CEECIND/03628/2017 CEECIND/04058/2018, European Union (EU) 847507, La Caixa Foundation ID 100010434, Portuguese Foundation for Science and Technology LCF/PR/HP17/52190003, PI20/01845, PI12/02688

Purpose. Previous studies have shown that melanoma cells produce excessive levels of cytokines, which have various biological roles during melanoma development. The aim of this study was to expand the profile of serum cytokines, chemokines, growth factors, and angiogenic factors that are associated with melanoma, to find more cytokines with abnormal concentrations in melanoma patients, to identify whether the level of cytokines correlated with prognostic variants, such as Breslow thickness and BRAF mutation, and, finally, to find out the cytokines that play important roles during melanoma development. Materials and Methods. Multiplex immunobead assay technology and 45-plex immunoassays ProcartaPlex (TM) kits were used to simultaneously compare the levels of cytokines, growth factors, angiogenic factors, and chemokines between the serum of healthy patients (n = 30) and those with melanoma (n = 72). Data were analyzed according to the clinical characteristics of the designated patient subgroups. Results. Compared to the control group, melanoma patients had higher levels of VEGF-A, PDGF-BB, IL-1RA, PIGF-1, IFN-gamma, TNF-alpha, MIP-1 alpha, and SCF, but lower levels of BDNF, SDF-1 alpha, MCP-1, Eotaxin, EGF, and IL-7. Furthermore, the levels of TNF-alpha (P=0.320, r = 0.019), IFN-gamma (P=0.311, r = 0.023), VEGF-A (P=0.014, r = 0.337), and BDNF (0.004, r = -0.391) showed a significant correlation with Breslow thickness. IL-7 was of lower levels in patients harboring BRAF mutants. Melanoma patients with high levels of MIP-1 alpha and MCP-1 showed the poorest overall survival. Conclusions. We found that the levels of VEGF-A and PDGF-BB in the serum of both primary and metastatic melanoma patients are elevated. TNF-alpha, IFN-gamma, and VEGF-A presented a positive correlation with Breslow thickness, whereas BDNF showed a negative association. MIP-1 alpha and MCP-1 correlated negatively with survival. In addition, lower levels of IL-7 were found in patients harboring BRAF mutants. These findings indicate that these cytokines may play critical roles in the progression of melanoma., Instituto de Salud Carlos II FISS PI 13/02119

This study was supported by grants PI17/02276 and PI20/01845 from Fondo de Investigaciones Sanitarias (Madrid, Spain) and by intramural funds of GENYO and FIBAO foundation (Granada, Spain). This study was also supported by the Novo Nordisk Fonden (NNF15OC0016932, VA) and Knud og Edith Eriksens Mindefond (VA) and Gigtforeningen (A2037, A3570, VA). JS and KB-K were supported by the grant No. 2016/21/B/NZ5/01901 from the National Science Centre (Poland). MGN was supported by a Spinoza grant from the Netherlands Organization for Scientific Research. YL was supported by an ERC Starting Grant (948207) and the Radboud University Medical Centre Hypatia Grant (2018) for Scientific Research. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript., We aimed to validate the association of 28 GWAS-identified genetic variants for response to TNF inhibitors (TNFi) in a discovery cohort of 1361 rheumatoid arthritis (RA) patients monitored in routine care and ascertained through the REPAIR consortium and DANBIO registry. We genotyped selected markers and evaluated their association with response to TNFi after 6 months of treatment according to the change in disease activity score 28 (DDAS28). Next, we confirmed the most interesting results through meta-analysis of our data with those from the DREAM cohort that included 706 RA patients treated with TNFi. The meta-analysis of the discovery cohort and DREAM registry including 2067 RA patients revealed an overall association of the LINC02549rs7767069 SNP with a lower improvement in DAS28 that remained significant after correction for multiple testing (perallele ORMeta=0.83, PMeta=0.000077; PHet=0.61). In addition, we found that each copy of the LRRC55rs717117G allele was significantly associated with lower improvement in DAS28 in rheumatoid factor (RF)-positive patients (per-allele ORMeta=0.67, P=0.00058; PHet=0.06) whereas an opposite but not significant effect was detected in RF-negative subjects (per-allele ORMeta=1.38, P=0.10; PHet=0.45; PInteraction=0.00028). Interestingly, although the identified associations did not survive multiple testing correction, the metaanalysis also showed overall and RF-specific associations for the MAFBrs6071980 and CNTN5rs1813443 SNPs with decreased changes in DAS28 (per-allele ORMeta_rs6071980 = 0.85, P=0.0059; PHet=0.63 and ORMeta_rs1813443_RF+=0.81, P=0.0059; PHet=0.69 and ORMeta_rs1813443_RF-=1.00, P=0.99; PHet=0.12; PInteraction=0.032). Mechanistically, we found that subjects carrying the LINC02549rs7767069T allele had significantly increased numbers of CD45RO+CD45RA+ T cells (P=0.000025) whereas carriers of the LINC02549rs7767069T/T genotype showed significantly increased levels of soluble scavengers CD5 and CD6 in serum (P=0.00037 and P=0.00041). In addition, carriers of the LRRC55rs717117G allele showed decreased production of IL6 after stimulation of PBMCs with B burgdorferi and E coli bacteria (P=0.00046 and P=0.00044), which suggested a reduced IL6-mediated anti-inflammatory effect of this marker to worsen the response to TNFi. In conclusion, this study confirmed the influence of the LINC02549 and LRRC55 loci to determine the response to TNFi in RA patients and suggested a weak effect of the MAFB and CNTN5 loci that need to be further investigated., Instituto de Salud Carlos III PI17/02276 PI20/01845, GENYO foundation (Granada, Spain), FIBAO foundation (Granada, Spain), Novo Nordisk Foundation NNF15OC0016932, Knud og Edith Eriksens Mindefond, Gigtforeningen A2037 A3570, National Science Centre, Poland 2016/21/B/NZ5/01901, Spinoza grant from the Netherlands Organization for Scientific Research, European Research Council (ERC) European Commission 948207, Radboud University Medical Centre Hypatia Grant (2018)

Here, we assessed whether 36 single nucleotide polymorphisms (SNPs) within the TNFSF4 and MAPKAPK2 loci influence the risk of developing invasive aspergillosis (IA). We conducted a two-stage case control study including 911 high-risk patients diagnosed with hematological malignancies that were ascertained through the aspBIOmics consortium. The meta-analysis of the discovery and replication populations revealed that carriers of the TNFSF4rs7526628T/T genotype had a significantly increased risk of developing IA (p = 0.00022). We also found that carriers of the TNFSF4rs7526628T allele showed decreased serum levels of TNFSF14 protein (p = 0.0027), and that their macrophages had a decreased fungicidal activity (p = 0.048). In addition, we observed that each copy of the MAPKAPK2rs12137965G allele increased the risk of IA by 60% (p = 0.0017), whereas each copy of the MAPKAPK2rs17013271T allele was estimated to decrease the risk of developing the disease (p = 0.0029). Mechanistically, we found that carriers of the risk MAPKAPK2rs12137965G allele showed increased numbers of CD38+IgM-IgD- plasmablasts in blood (p = 0.00086), whereas those harboring two copies of the allele had decreased serum concentrations of thymic stromal lymphopoietin (p = 0.00097). Finally, we also found that carriers of the protective MAPKAPK2rs17013271T allele had decreased numbers of CD27-IgM-IgD- B cells (p = 0.00087) and significantly lower numbers of CD14+ and CD14+CD16- cells (p = 0.00018 and 0.00023). Altogether, these results suggest a role of the TNFSF4 and MAPKAPK2 genes in determining IA risk.

We thank all participants who have agreed to participate in this study. Authors also thank Maria Dolores Casares, Angeles Molina, Carmen Oloriz for the collection of Spanish samples and Hans Jurgen Hoffmann, Marianne Thomsen, Vibeke Ostergaard Thomsen, Malene Rohr Andersen, Lise Lotte B. Laursen, Helle Jorgensen, Ram Benny Christian Dessau, Niels Steen Krogh, Ulla Vogel, Paal Skytt Andersen, Ivan Brandslund, Steffen Bank, Frederik Trier Moller, Nikolai Toft and Niels Moller Andersen for the participation in collection and purification of Danish samples. We also thank the Danish Departments of Rheumatology for their implication in the collection of clinical data from RA patients included in the DANBIO cohort and the Danish Rheumatologic Biobank. Likewise, we would like to thank Teun van Herwaarden for steroid hormone measurements in serum samples from subjects ascertained through the HFGP initiative. This work was partially supported by intramural funds of GENYO and FIBAO foundation (Granada, Spain); Novo Nordisk Fonden (NNF15OC0016932, VA); and Knud og Edith Eriksens Mindefond (VA) and Gigtforeningen (A2037, A3570, VA). MGN was supported by a Spinoza grant from the Netherlands Organization for Scientific Research., All data used in this project have been meticulously cataloged and archived in the BBMRI-NL data infrastructure (https://hfgp.bbmri.nl/) using the MOLGENIS open source platform for scientific data45. This allows flexible data querying and download, including sufficiently rich metadata and interfaces for machine processing (R statistics, REST API) and using FAIR principles to optimize Findability, Accessibility, Interoperability and Reusability46. Genetic data from the discovery and DANBIO populations can be accessed at ftp.genyo.es and data from the DREAM registry are available at https://www.synapse.org/#!Synapse:syn3280809/wiki/194735 and https://www. synapse.org/#!Synapse:syn3280809/wiki/194736., Supplementary information is available for this paper at https://doi.org/10.1038/s41598-020-61331-5., This study sought to evaluate the association of 28 single nucleotide polymorphisms (SNPs) within NFKB and inflammasome pathway genes with the risk of rheumatoid arthritis (RA) and response to TNF inhibitors (TNFi). We conducted a case-control study in a European population of 1194 RA patients and 1328 healthy controls. The association of potentially interesting markers was validated with data from the DANBIO (695 RA patients and 978 healthy controls) and DREAM (882 RA patients) registries. The meta-analysis of our data with those from the DANBIO registry confirmed that anti-citrullinated protein antibodies (ACPA)-positive subjects carrying the NFKB2rs11574851T allele had a significantly increased risk of developing RA (PMeta_ACPA + = 0.0006) whereas no significant effect was found in ACPA-negative individuals (PMeta_ACPA− = 0.35). An ACPA-stratified haplotype analysis including both cohorts (n = 4210) confirmed that ACPA-positive subjects carrying the NFKB2TT haplotype had an increased risk of RA (OR = 1.39, P = 0.0042) whereas no effect was found in ACPA-negative subjects (OR = 1.04, P = 0.82). The meta-analysis of our data with those from the DANBIO and DREAM registries also revealed a suggestive association of the NFKB2rs1056890 SNP with larger changes in DAS28 (OR = 1.18, P = 0.007). Functional experiments showed that peripheral blood mononuclear cells from carriers of the NFKB2rs1005044C allele (in LD with the rs1056890, r2 = 1.00) showed increased production of IL10 after stimulation with LPS (P = 0.0026). These results provide first evidence of a role of the NFKB2 locus in modulating the risk of RA in an ACPA-dependent manner and suggest its implication in determining the response to TNFi. Additional studies are now warranted to further validate these findings., GENYO, FIBAO foundation (Granada, Spain), Novo Nordisk Foundation NNF15OC0016932, Knud og Edith Eriksens Mindefond A2037 A3570, Gigtforeningen A2037 A3570, Spinoza grant from the Netherlands Organization for Scientific Research

We are deeply grateful to the study participants. We also thank Astella Pharma Inc. and Mrs. Consuelo González Moreno (AML survivor) for supporting this work., The purpose of this study was to conduct a two-stage case control association study including 654 acute myeloid leukaemia (AML) patients and 3477 controls ascertained through the NuCLEAR consortium to evaluate the effect of 27 immune-related single nucleotide polymorphisms (SNPs) on AML risk. In a pooled analysis of cohort studies, we found that carriers of the IL13rs1295686A/A genotype had an increased risk of AML (PCorr = 0.0144) whereas carriers of the VEGFArs25648T allele had a decreased risk of developing the disease (PCorr = 0.00086). In addition, we found an association of the IL8rs2227307 SNP with a decreased risk of developing AML that remained marginally significant after multiple testing (PCorr = 0.072). Functional experiments suggested that the effect of the IL13rs1295686 SNP on AML risk might be explained by its role in regulating IL1Ra secretion that modulates AML blast proliferation. Likewise, the protective effect of the IL8rs2227307 SNP might be mediated by TLR2-mediated immune responses that affect AML blast viability, proliferation and chemorresistance. Despite the potential interest of these results, additional functional studies are still warranted to unravel the mechanisms by which these variants modulate the risk of AML. These findings suggested that IL13, VEGFA and IL8 SNPs play a role in modulating AML risk., Astella Pharma Inc., Instituto de Salud Carlos III PI12/02688 PI17/02276, Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) NORTE-01-0145-FEDER-000013, Portuguese Foundation for Science and Technology CEECIND/04601/2017 CEECIND/03628/2017, Astellas Pharmaceuticals

Here, we assessed whether 41 SNPs within steroid hormone genes associated with erosive disease. The most relevant finding was the rheumatoid factor (RF)-specific effect of the CYP1B1, CYP2C9, ESR2, FcγR3A, and SHBG SNPs to modulate the risk of bone erosions (P = 0.004, 0.0007, 0.0002, 0.013 and 0.015) that was confirmed through meta-analysis of our data with those from the DREAM registry (P = 0.000081, 0.0022, 0.00074, 0.0067 and 0.0087, respectively). Mechanistically, we also found a gender-specific correlation of the CYP2C9rs1799853T/T genotype with serum vitamin D3 levels (P = 0.00085) and a modest effect on IL1β levels after stimulation of PBMCs or blood with LPS and PHA (P = 0.0057 and P = 0.0058). An overall haplotype analysis also showed an association of 3 ESR1 haplotypes with a reduced risk of erosive arthritis (P = 0.009, P = 0.002, and P = 0.002). Furthermore, we observed that the ESR2, ESR1 and FcγR3A SNPs influenced the immune response after stimulation of PBMCs or macrophages with LPS or Pam3Cys (P = 0.002, 0.0008, 0.0011 and 1.97•10−7). Finally, we found that a model built with steroid hormone-related SNPs significantly improved the prediction of erosive disease in seropositive patients (PRF+ = 2.46•10−8) whereas no prediction was detected in seronegative patients (PRF− = 0.36). Although the predictive ability of the model was substantially lower in the replication population (PRF+ = 0.014), we could confirm that CYP1B1 and CYP2C9 SNPs help to predict erosive disease in seropositive patients. These results are the first to suggest a RF-specific association of steroid hormone-related polymorphisms with erosive disease.

BACKGROUND Alopecia areata (AA) is a skin disease that produces hair loss in patches of skin. The underlying mechanism of AA is a loss of immune privilege of hair follicles, which are then attacked by natural killer (NK) cells. A previous genome-wide association study linked single nucleotide polymorphisms of the protein MHC class I chain-related A (MICA) to this disease. MICA is the ligand for the activating receptor NKG2D, expressed mainly by NK cells and CD8+ cytotoxic T cells. As the aforementioned study did not include short tandem repeats (STRs) of MICA, we decided to study these in relation to AA. AIM To study the association of STRs with AA, alongside that of human leucocyte antigen (HLA) locus B, which is closely linked to MICA. METHODS DNA amplicon size analysis was carried out, and HLA-B locus genomic typing was performed by PCR-sequence-specific oligonucleotide analysis. RESULTS We observed an association between AA and both MICA*009 and HLA-B14; associations were also observed between HLA-B alleles and MICA alleles, which have both been previously found to be connected with AA, but never studied together. CONCLUSIONS We conclude that it is important to study HLA-B and MICA together to avoid the influence of their association in experiments in which they are investigated separately.

Background:The interplay between genetics and drug response in rheumatoid arthritis (RA) has shown that response to biologics varies between individuals and that a large proportion of patients show no clinical improvement (Plenge and Bridges, 2011). Despite the disappointing scenario, to date, only a few genetic markers have been consistently identified and we are far from being able to optimize drug dosing or prioritize drug combinations based on genetic findings.Objectives:With this background, we sought to validate the association of GWAS-identified variants for response to TNF inhibitors (TNFi) in a two-stage case control association study and to shed some light into the functional role of the most interesting markers.Methods:The discovery population consisted of 1361 RA patients ascertained through the REPAIR consortium and the DANBIO registry. RA patients fulfilled the 1987 revised American College of Rheumatology (ACR) and the ACR/EULAR 2010 classification criteria. The validation cohort included 706 Dutch RA patients from the DREAM registry. The study followed the Declaration of Helsinki and study participants gave their written informed consent to participate in the study, which was approved by the ethical review committee of participant institutions. Twenty-seven single-nucleotide polymorphisms (SNPs) were selected through a literature search of relevant GWAS. Linear regression analysis adjusted for age, sex and country of origin was used to determine the association between GWAS-identified SNPs and changes in DAS28 (ΔDAS28) after 3 or 6 months of treatment. The meta-analysis of both populations was performed using a fixed effect model. Correction for multiple testing was performed using the Bonferroni method but also considering the number of inheritance models tested (P=0.0009). To assess the role of the most interesting markers in modulating immune responses, stimulation experiments in whole blood, peripheral mononuclear cells (PBMCs) and monocyte-derived macrophages using a large number of pathogens and microbiome bacteria were performed in 408 subjects from the Human Functional Genomic Project cohort. We also evaluated the correlation of these SNPs with plasmatic levels of 108 inflammatory proteins, 7 serum steroid hormones and counts of 91 blood-derived immune cell populations.Results:The meta-analysis of the discovery cohort and DREAM registry including 2067 RA patients treated with TNFi revealed an overall association of the LINC02549rs7767069 SNP with a decreased drop in DAS28 that remained significant after correction for multiple testing (per-allele ORMeta=0.83, PMeta=0.000077; PHet=0.61). In addition, the meta-analysis of these large cohorts showed that each copy of the LARRC55rs717117G allele significantly decreased the drop in DAS28 in RF-positive patients (per-allele ORMeta=0.67, P=0.00058; PHet=0.06) whereas an opposite but not significant effect was found in RF-negative subjects (per-allele ORMeta=1.38, P=0.10; PInteraction=0.00028; PHet=0.45). Interestingly, the meta-analysis also showed potentially interesting but not statistically significant overall and RF-specific associations for the MAFBrs6071980 and CNTN5rs1813443 SNPs with ΔDAS28 (per-allele ORMeta_rs6071980=0.84, P=0.0059; PHet=0.63 and ORMeta_rs1813443_RF+=0.81, P=0.0059; PHet=0.69 and ORMeta_rs1813443_RF-=1.00, P=0.99; PHet=0.12; PInteraction=0.032). Although analysis of functional data is ongoing, so far, we found that carriers of the LARRC55rs717117G allele showed decreased levels of IL6 after stimulation of PBMCs with Borrelia burgdorferi and Escherichia Coli bacteria (P=0.00046 and 0.00044), which suggested a reduced IL6-mediated anti-inflammatory effect of this marker to worsen the response to TNFi.Conclusion:This study confirmed the influence of the LINC02549 and LARRC55 loci to determine the response to TNFi in RA patients and a weak effect of the MAFB and CNTN5 loci that needs to be further investigated.References:[1]Plenge RM et al 2011. Arthritis Rheum 63, 590-3.ACKNOWLEDGEMENTS:We thank all participants who have agreed to participate in this study. Authors also thank María Dolores Casares, Ángeles Molina, Carmen Oloriz for the collection of Spanish samples and Hans Jurgen Hoffmann, Marianne Thomsen, Vibeke Østergaard Thomsen, Malene Rohr Andersen, Lise Lotte B. Laursen, Helle Jørgensen, Ram Benny Christian Dessau, Niels Steen Krogh, Ulla Vogel, Paal Skytt Andersen, Ivan Brandslund, Steffen Bank, Frederik Trier Møller, Nikolai Toft and Niels Møller Andersen for the participation in collection and purification of Danish samples. We also thank the Danish Departments of Rheumatology for their implication in the collection of clinical data from RA patients included in the DANBIO cohort and the Danish Rheumatologic Biobank. Likewise, we would like to thank Teun van Herwaarden for steroid hormone measurements in serum samples from subjects ascertained through the HFGP initiative.Disclosure of Interests:None declared

To investigate the proportion of asymptomatic infection among blood donors in a region endemic for Leishmania; and to ascertain epidemiological and genetic factors associated with this condition. We studied 1260 blood donors in the Province of Granada in the Southern Spain. After obtaining informed consent in each participant, a poll about habits, housing and contact with animals were carried out. Blood samples were obtained for determining antileishmanial antibodies and a PCR assay. HLA typing was performed in a randomly sample among the donors with positive serology. We have found that L. infantum antibodies were present in 7.9% of blood donors and DNA in blood was detected in 2.5% of donors. There was no concordance between both determinations, except in one patient. Taking into consideration both techniques, 129 participants were considered to have asymptomatic Leishmania infection. No participant in this study developed clinical leishmaniasis during a follow-up period of 2 years. HLA were typed in 51 donors. Asymptomatic Leishmania infection might be associated with certain HLA antigens. A multivariate analysis was done with the variables obtained through the participants’ interview. The contact with livestock (goats, pigs, and sheep), but not dogs, either at home or in the environment, was significantly and independently associated with asymptomatic leishmania infection. Asymptomatic leishmanial infection among blood donors is frequent in the Granada Province, south of Spain. The presence of livestock in this region is related to this infection, perhaps influencing vector density of this disease. Some HLA genes might be associated with asymptomatic leishmanial state.

BACKGROUND/OBJECTIVES The aetiology of frontal fibrosing alopecia is unknown, and its genetic aspect remains uncharacterised. The aim of this report is to elucidate if major histocompatibility complex is associated with familial frontal fibrosing alopecia. METHODS A case-control study was performed of 13 patients with frontal fibrosing alopecia belonging to six families. Their human leukocyte antigen profiles were compared to the data of 636 healthy controls without frontal fibrosing alopecia. Patients underwent high-resolution genomic typing for human leukocyte antigen class I and II loci by PCR-SSO for Luminex. In addition, CYP21A2 gene (major histocompatibility complex class III) mutations were detected by PCR-SSO on strips. RESULTS 61.5% of patients shared CYP21A2 gene p.V281L linked to the F16A human leukocyte antigen class I haplotype (HLA-A*33:01; B*14:02; C*08:02; Pc

The aetiology of frontal fibrosing alopecia is unknown, and its genetic aspect remains uncharacterised. The aim of this report is to elucidate if major histocompatibility complex is associated with familial frontal fibrosing alopecia.A case-control study was performed of 13 patients with frontal fibrosing alopecia belonging to six families. Their human leukocyte antigen profiles were compared to the data of 636 healthy controls without frontal fibrosing alopecia. Patients underwent high-resolution genomic typing for human leukocyte antigen class I and II loci by PCR-SSO for Luminex. In addition, CYP21A2 gene (major histocompatibility complex class III) mutations were detected by PCR-SSO on strips.61.5% of patients shared CYP21A2 gene p.V281L linked to the F16A human leukocyte antigen class I haplotype (HLA-A*33:01; B*14:02; C*08:02; Pc 0.000001). The patients F16A-negative shared other human leukocyte antigen class I haplotypes: Y16A (3/13) and S26 (2/13).CYP21A2 gene p.V281L mutation can be used as a genetic marker for susceptibility to familial frontal fibrosing alopecia. Both the linkage of the mutation to F16A and the fact that F16A-negative patients share other human leukocyte antigen class I haplotype, point to an antigen-driven mechanism in susceptible patients with these haplotypes.

Background/objectives Frontal fibrosing alopecia (FFA) is a scarring alopecia whose prevalence is increasing. The pathogenesis of this disease is not well known. Genetic, environmental, hormonal and autoimmunity related factors have been considered; however, only a few cases of familial frontal fibrosing alopecia have been reported. Material and methods A cross-sectional study was performed at University Hospital in Granada (Spain). Twenty patients with frontal fibrosing alopecia belonging to nine different families were included, and clinical and dermoscopic features were analysed. Results Overall, 90% of the patients studied were women (mean age 61.4 years). About 50% of the patients had grade II frontal fibrosing alopecia at the time of diagnosis, whilst 35% had grades III or V. Mean recession was 2.83 cm in the frontal area and 1.99 cm in the temporo-parietal area. Daughters presented a shorter recession area and earlier debut of the disease than mothers. Androgenetic alopecia was found in only two patients (10%). The dermoscopic signs most commonly found were perifollicular erythema (85%), hyperkeratosis (85%), and absence of vellus hair in the hairline (78.9%). Conclusion This study adds to the growing evidence that there is a genetic component to frontal fibrosing alopecia. The clinical pattern of frontal fibrosing alopecia was not different from that found in non-familial cases, but the debut of the disease in daughters of mothers with frontal fibrosing alopecia may be earlier.

Background Although the etiology of rheumatoid arthritis (RA) remains unclear, there are evidences suggesting a role of sex steroid hormones in determining the onset and progression of the disease. Objectives The aim of this study was to evaluate whether 47 single nucleotide polymorphisms (SNPs) in steroid hormone-related genes are associated with the risk of RA and anti-TNF drug response. Methods We conducted a case-control study in 3 European populations including 2936 RA patients and 2197 healthy controls. Of those a total of 1985 RA patients were treated with anti-TNF blockers. The association of potentially interesting markers with RA risk or drug response in the discovery population was validated through meta-analysis with data from DREAM and DANBIO registries. Results were corrected for multiple testing (Meff method; p=0.0011). The value of steroid hormone-related variants for prediction of anti-TNF drug response was also assessed using stepwise logistic regression analyses. The area under the curve (AUC) of a receiver operating characteristic (ROC) curve analysis and a −2 log likelihood ratio (LR) test were used to assess whether the genetic model fitted significantly better the data compared to the reference model. A randomization test (50.000 iterations) was run to confirm the consistency of the results. Results Although none of the selected variants played a relevant role in modulating RA risk, the meta-analysis of the linear regression data with those from the DREAM and DANBIO registries showed a significant correlation of the CYP3A4 rs11773597 and CYP2C9 rs1799853 variants with changes in DAS28 after the administration of anti-TNF drugs (p=0.00074, and p=0.006). An overall haplotype analysis also showed that the ESR2 GGG haplotype significantly associated with a reduced chance of having poor response to anti-TNF drugs (p=0.0009). Finally, a ROC curve analysis confirmed that a model built with 8 steroid hormone-related variants significantly improved the ability to predict drug response compared with the reference model including demographic and clinical variables (AUC=0.633 vs. 0.556; P LRtest =1.52·10–6). Conclusions These data suggest that steroid hormone-related genes play a role in determining the response to anti-TNF drugs. Disclosure of Interest L. Canet: None declared, J. Sanchez-Maldonado: None declared, A. Rodriguez-Ramos: None declared, C. Lupianez: None declared, H. Canhao: None declared, M. Martinez-Bueno: None declared, A. Escudero: None declared, J. Segura-Catena: None declared, S. Sorensen: None declared, M. Hetland: None declared, M. Soto-Pino: None declared, M. Ferrer: None declared, A. Garcia: None declared, B. Glintborg: None declared, I. Filipescu: None declared, E. Perez-Pampin: None declared, A. Gonzalez-Utrilla: None declared, M. Lopez-Nevot: None declared, P. Conesa-Zamora: None declared, A. den Broeder: None declared, S. da Vita: None declared, S. Hove Jacobsen: None declared, E. Collantes: None declared, L. Quartuccio: None declared, J. Fonseca: None declared, M. Coenen: None declared, V. Andersen Consultant for: VA has received compensation for consultancy and for being a member of an advisory board from MSD (Merck) and Janssen, R. Caliz-Caliz: None declared, J. Sainz: None declared

Background Rheumatoid arthritis (RA) is a chronic autoimmune disease that arises as a result of the interaction between genetic and environmental factors. A growing body of research suggests that genetic variants within immune-related genes can influence the risk of developing the disease and affect drug response. Materials and methods To test this hypothesis, we carried out a comprehensive two-stage case–control study in a White population of 1239 White RA patients and 1229 healthy controls to investigate whether 49 single nucleotide polymorphisms within or near 17 immune-related genes modulate the risk of developing RA and antitumor necrosis factor (anti-TNF) drug response. Results Logistic regression analyses showed that carriers of the IL4rs2070874T and IL4rs2243250T and IL8RBrs1126580A alleles or the IL8RBrs2230054C/C genotype had a significantly increased risk of developing RA [odds ratio (OR)=1.37, 95% confidence interval (CI) 1.13–1.67, P=0.0016; OR=1.24, 95% CI 1.03–1.49, P=0.020; OR=1.23, 95% CI 1.08–1.41, P=0.002 and OR=1.19, 95% CI 1.04–1.36, P=0.01, respectively]. The association of the IL4 variants was further supported by a meta-analysis including 7150 individuals (P =0.0010), whereas the involvement of the IL8RB locus in determining the susceptibility to RA was also supported by gene–gene interaction analyses that identified significant two-locus and three-locus interaction models including IL8RB variants that act synergistically to increase the risk of the disease (P=0.014 and 0.018). Interestingly, we also found that patients harbouring the IFNGrs2069705C allele showed a significantly better response to anti-TNF drugs than those patients carrying the wild-type allele (P=0.0075). Conclusions Our data suggest that IL4 and IL8RB loci may have a small-effect genetic impact on the risk of developing RA, whereas IFNG might be involved in modulating the response to anti-TNF drugs.

© Springer Nature Limited 2018, The aim of this case-control study was to evaluate whether 47 single-nucleotide polymorphisms (SNPs) in steroid hormone-related genes are associated with the risk of RA and anti-TNF drug response. We conducted a case-control study in 3 European populations including 2936 RA patients and 2197 healthy controls. Of those, a total of 1985 RA patients were treated with anti-TNF blockers. The association of potentially interesting markers in the discovery population was validated through meta-analysis with data from DREAM and DANBIO registries. Although none of the selected variants had a relevant role in modulating RA risk, the meta-analysis of the linear regression data with those from the DREAM and DANBIO registries showed a significant correlation of the CYP3A4rs11773597 and CYP2C9rs1799853 variants with changes in DAS28 after the administration of anti-TNF drugs (P = 0.00074 and P = 0.006, respectively). An overall haplotype analysis also showed that the ESR2GGG haplotype significantly associated with a reduced chance of having poor response to anti-TNF drugs (P = 0.0009). Finally, a ROC curve analysis confirmed that a model built with eight steroid hormone-related variants significantly improved the ability to predict drug response compared with the reference model including demographic and clinical variables (AUC = 0.633 vs. AUC = 0.556; PLR_test = 1.52 × 10-6). These data together with those reporting that the CYP3A4 and ESR2 SNPs correlate with the expression of TRIM4 and ESR2 mRNAs in PBMCs (ranging from P = 1.98 × 10-6 to P = 2.0 × 10-35), and that the CYP2C9rs1799853 SNP modulates the efficiency of multiple drugs, suggest that steroid hormone-related genes may have a role in determining the response to anti-TNF drugs.KEY POINTS• Polymorphisms within the CYP3A4 and CYP2C9 loci correlate with changes in DAS28 after treatment with anti-TNF drugs.• A haplotype including eQTL SNPs within the ESR2 gene associates with better response to anti-TNF drugs.• A genetic model built with eight steroid hormone-related variants significantly improved the ability to predict drug response., This work was supported by grants from FIBAO foundation (Granada, Spain), Novo Nordisk Fonden (NNF15OC0016932), Knud og Edith Eriksens Mindefond and Gigtforeningen (A2037, A3570).

Toll-like receptors trigger the innate immune response by activating various cell types such us macrophages and lymphocytes. We genotyped SNV of TLR3, TRL7, TLR8 and TLR10 in 863 Spanish and 150 Italian patients with Meniere's disease (MD) and 1,013 controls by using Taqman assays. Real-Time qPCR was used to measure the expression level of TLR10 in peripheral blood leukocytes. The overall dataset showed that the C allele and the CC genotype of rs11096955 in TLR10 gene were more commonly observed in controls than patients (corrected p = 1 × 10(-3), OR = 0.68 [95 % confidence interval, 0.54-0.84] for CC genotype; corrected p = 1.5 × 10(-5), OR = 0.75 [0.66-0.85] for allele C). Moreover, the CC genotype was more frequent in patients with uni- (19 %) than bilateral sensorineural hearing loss (SNHL) (13 %). Logistic regression demonstrated that the time since the onset of MD, Tumarkin crises, hearing stage and rs11096955 were independent factors influencing the risk of bilateral SNHL. In addition, rs11096955 influenced hearing loss progression in patients with bilateral MD. No change in expression of TLR10 was observed according to CC, CA or AA genotypes. Our data suggest that allelic variants of TLR10 gene may influence the susceptibility and time-course of hearing loss of MD in the European population.

Variability in acute immune response genes could determine susceptibility or prognosis for Ménière's disease (MD). The cytokines tumor necrosis factor α (TNFα), macrophage migration inhibitory factor (MIF) and interferon γ (INFγ) are proinflammatory cytokines of the innate immune response. These cytokines mediate inflammation and have been previously associated with the inflammatory process in several autoimmune diseases. We investigated the association between functional allelic variants of MIF (rs35688089), IFNG (rs2234688) and TNFA (rs1800629) in patients with MD. In addition to testing these variants for an association with disease, we also tested for an association with clinical aspects of disease progression, such as persistence of vertigo and the sensorineural hearing loss. A total of 580 patients with diagnosis of definite MD, according to the diagnostic scale of the American Academy of Otolaryngology-Head and Neck Surgery, and 552 healthy controls were included. DNA samples from a set of 291 American patients were used to confirm the results obtained in the MIF gene in our Spanish cohort. Although we found a significant association with the allele containing five repeats of CATT within the MIF gene in patients with MD in the Spanish cohort [corrected p = 0.008, OR = 0.69 (95 % CI, 0.54-0.88)], this finding could not be replicated in the American set. Moreover, no genetic associations for variants in either the TNFA or IFNG genes and MD were found. These results support the conclusion that functional variants of MIF, INFG, and TFNA genes are not associated with disease susceptibility or hearing loss progression in patients with MD.

To evaluate prospectively the usefulness of the routine determination of BRAF(T1799A) mutation on thyroid fine-needle aspiration biopsy (FNAB) to detect cytopathology false negative papillary thyroid carcinomas (PTC) and, therefore, as a tool to improve the sensitivity of the preoperative cytopathological diagnosis of PTC in thyroid nodules.FNAB is the most reliable diagnostic test to discriminate between malignant and benign thyroid nodules, but nondiagnostic results remain a clinical management dilemma. BRAF(T1799A) mutation is the most prevalent genetic alteration in thyroid cancers and is specific for PTC, characteristics that make it the most potentially helpful genetic tool to improve the diagnostic accuracy of FNAB.An exhaustive recruitment of all patients subjected to thyroid FNAB in our institution during 4 years was performed. BRAF(T1799A) mutation was determined on thyroid FNAB specimens by PCR and restriction fragment length polymorphism, plus direct sequencing in positive samples.BRAF(T1799A) mutation on FNAB detected 47.2% (17/36) of PTC cases. It confirmed preoperatively 45.5% (5/11) of the PTC cases in the indeterminate category and decreased the rate of cytopathology false-negatives in 33.3% (6/18), improving the combined (BRAF(T1799A) mutation + cytopathological analysis) sensitivity of the detection of PTC on FNAB in 16.7%.BRAF(T1799A) mutation improves the diagnosis of PTC on FNAB, mainly because of the detection of cytopathology false-negatives, and it can be helpful in the routine analysis of thyroid nodules, especially in clinical settings with moderate sensitivity to detect PTC on FNAB.

ObjectivesSystemic lupus erythematosus (SLE) is a chronic multisystem genetically complex autoimmune disease characterised by the production of autoantibodies to nuclear and cellular antigens, tissue inflammation and organ damage. Genome-wide association studies have shown that variants within the major histocompatibility complex (MHC) region on chromosome 6 confer the greatest genetic risk for SLE in European and Chinese populations. However, the causal variants remain elusive due to tight linkage disequilibrium across disease-associated MHC haplotypes, the highly polymorphic nature of many MHC genes and the heterogeneity of the SLE phenotype.MethodsA high-density case-control single nucleotide polymorphism (SNP) study of the MHC region was undertaken in SLE cohorts of Spanish and Filipino ancestry using a custom Illumina chip in order to fine-map association signals in these haplotypically diverse populations. In addition, comparative analyses were performed between these two datasets and a northern European UK SLE cohort. A total of 1433 cases and 1458 matched controls were examined.ResultsUsing this transancestral SNP mapping approach, novel independent loci were identified within the MHC region in UK, Spanish and Filipino patients with SLE with some evidence of interaction. These loci include HLA-DPB1, HLA-G and MSH5 which are independent of each other and HLA-DRB1 alleles. Furthermore, the established SLE-associated HLA-DRB1*15 signal was refined to an interval encompassing HLA-DRB1 and HLA-DQA1. Increased frequencies of MHC region risk alleles and haplotypes were found in the Filipino population compared with Europeans, suggesting that the greater disease burden in non-European SLE may be due in part to this phenomenon.ConclusionThese data highlight the usefulness of mapping disease susceptibility loci using a transancestral approach, particularly in a region as complex as the MHC, and offer a springboard for further fine-mapping, resequencing and transcriptomic analysis.

Objective. To determine the role of Class II HLAs in SSc patients from Italy and Spain and in SSc patients of Caucasian ancestry. Methods. Nine hundred and forty-four SSc patients (Italy 392 patients; Spain 452 patients) and 1320 ethnically matched healthy controls (Italy 398 patients; Spain 922 patients) were genotyped up to the fourth digit by PCR with sequence-specific oligonucleotides for HLA-DRB1, DQA1 and DQB1 loci. Patients included 390 ACA-positive and 254 anti-topo I-positive subjects. Associations between SSc or SScspecific antibodies and HLA alleles or HLA haplotypes were sought via the chi-square test after 10 000-fold permutation testing. A meta-analysis including this study cohort and other Caucasoids samples was also conducted. Results. In both the cohorts, the strongest association was observed between the HLA-DRB1*1104 allele and SSc or anti-topo I antibodies. The HLA-DRB1*1104 -DQA1*0501 -DQB1*0301 haplotype was overrepresented in Italian [odds ratio (OR) = 2.069, 95% asymptotic CIs (CI95) 1.486, 2.881; P < 0.001] and in Spanish patients (OR = 6.707, CI95 3.974, 11.319; P < 0.001) as well as in anti-topo-positive patients: Italy (OR = 2.642, CI95 1.78, 3.924; P < 0.001) and Spain (OR = 20.625, CI95 11.536, 36.876; P < 0.001). In both the populations we also identified an additional risk allele (HLA-DQB1*03) and a protective allele (HLADQB1*0501) in anti-topo-positive patients. The meta-analysis showed different statistically significant associations, the most interesting being the differential association between HLA-DRB1*01 alleles and ACAs (OR = 1.724, CI95 1.482, 2.005; P < 0.001) or topo I antibodies (OR = 0.5, CI95 0.384, 0.651; P < 0.001). Conclusions. We describe multiple robust associations between SSc and HLA Class II antigens in Caucasoids that may help to understand the genetic architecture of SSc.

Resumen Antecedentes y objetivo La mutacion BRAFT1799A se ha relacionado con caracteristicas tumorales de mas agresividad, recidiva tumoral y persistencia de carcinoma papilar de tiroides (CPT), aunque no todos los estudios apoyan esta asociacion. En este, se analiza la asociacion entre la presencia de la mutacion BRAFT1799A en el tumor primario de pacientes con CPT y las caracteristicas clinicopatologicas de riesgo, recidiva y persistencia tumoral. Pacientes, material y metodo Hemos seguido a 97 pacientes intervenidos de CPT durante una mediana de 64,1 meses. La mutacion BRAFT1799A se determino en acido desoxirribonucleico procedente de muestras de la tiroidectomia inicial mediante amplificacion por PCR del exon 15 del gen braf y analisis de los fragmentos de restriccion con la enzima TspRI. Los casos positivos fueron confirmados por secuenciacion. La asociacion estadistica entre la mutacion BRAFT1799A y las diferentes variables se estudio mediante los correspondientes tests de contraste de hipotesis mas regresion logistica. Resultados El 46,4% de los pacientes eran positivos para la mutacion BRAFT1799A. Tras analisis bivariante y multivariante, la mutacion BRAFT1799A solo se asociaba con edad superior a 60 anos (odds ratio [OR] = 5,5; intervalo de confianza [IC] del 95%, 1,4-21,9; p = 0,019) y tamano de 1 cm o superior (OR = 3,6; IC del 95%, 1,2-10,3; p = 0,016). No se asociaba con subtipos histologicos, metastasis, recidiva, necesidad de nuevos tratamientos ablativos con I131 o de otras intervenciones quirurgicas debidas a la aparicion de metastasis o persistencia de enfermedad al final del seguimiento. Conclusiones La mutacion BRAFT1799A esta asociada a edad superior a 60 anos y tamano tumoral de 1 cm o mayor, pero no con otras caracteristicas clinicopatologicas, recidiva tumoral o persistencia de enfermedad.

Background and objective The BRAF T1799A mutation is reported to be associated with aggressive, persistent, and recurrent tumor in patients with papillary thyroid carcinoma (PTC). The association of the BRAF T1799A mutation in the primary tumor with the clinicopathological characteristics of PTC was analyzed. Patients, materials, and methods Ninety-seven PTC patients were followed up for a median of 64.1 months. The BRAF T1799A mutation was analyzed in DNA from initial thyroidectomy biopsies by PCR amplification and restriction fragment length polymorphism using the TspRI enzyme. Positive results were confirmed by DNA sequencing. The statistical association of the BRAF T1799A mutation and clinicopathological characteristics was analyzed using the relevant hypothesis tests and logistic regression. Results The BRAF T1799A mutation was found in 46.4% of patients. Bivariate and multivariate analyses showed the mutation to be only associated with age over 60 years (odds ratio [OR] = 5.5; 95% confidence interval [CI], 1.4–21.9; p = 0.019) and to a tumor size of 1 cm or greater (OR = 3.6, 95% CI, 1.2–10.3; p = 0.016). The mutation was not associated with histological subtype, metastasis, recurrence, more aggressive treatments (131I ablation therapy or other surgery), or PTC persistence at the end of follow-up. Conclusions The BRAF T1799A mutation is associated with age over 60 and a tumor size of 1 cm or greater, but not with other clinicopathological characteristics, tumor recurrence, or PTC persistence.

Item does not contain fulltext BACKGROUND: The Fc receptors II and III (FcgR2a, and FcgR3a) play a crucial role in the regulation of the immune response. The FcgR2a*519GG and FcgR3a*559CC genotypes have been associated with several autoimmune diseases including systemic lupus erythematosus, rheumatoid arthritis, nephritis, and possibly to type I diabetes, and celiac disease. In a large multicenter, two-stage study of 6570 people, we tested whether the FcgR2a and FcgR3a genes were also involved in inflammatory bowel disease (IBD), which includes Crohn's disease (CD) and ulcerative colitis (UC). METHODS: We genotyped the FcgR2a*A519G and FcgR3a*A559C functional variants in 4205 IBD patients in six well-phenotyped Caucasian IBD cohorts and 2365 ethnically matched controls recruited from the Netherlands, Spain, and New Zealand. RESULTS: In the initial Dutch study we found a significant association of FcgR2a genotypes with IBD (P-genotype = 0.02); while the FcgR2a*519GG was more common in controls (23%) than in IBD patients (18%; odds ratio [OR] = 0.75; 95% confidence interval [CI] 0.61-0.92; P = 0.004). This association was corroborated by a combined analysis across all the study populations (Mantel-Haenszel [MH] OR = 0.84; 0.74-0.95; P = 0.005) in the next stage. The Fcgr2a*GG genotype was associated with both UC (MH-OR = 0.84; 0.72-0.97; P = 0.01) and CD (MH-OR = 0.84; 0.73-0.97; P = 0.01), suggesting that this genotype confers a protective effect against IBD. There was no association of FcgR3a*A559C genotypes with IBD, CD, or UC in any of the three studied populations. CONCLUSIONS: The FcgR2a*519G functional variant was associated with IBD and reduced susceptibility to UC and to CD in Caucasians. There was no association between FcgR3a*5A559C and IBD, CD or UC. 01 december 2010

Primary congenital glaucoma (PCG) is a genetically heterogeneous autosomal recessive disorder, which is an important cause of blindness in childhood. The first known gene, CYP1B1, accounts for a variable proportion of cases in most populations. A second gene, LTBP2, was recently reported in association with a syndrome, in which glaucoma is secondary to lens dislocation. We report on the molecular and clinical profile of 34 families diagnosed as PCG, all originating from the Roma/Gypsy founder population. Comprehensive sequencing analysis revealed a level of heterogeneity unusual for this population, with five CYP1B1 and one ancestral LTBP2 mutation accounting for ∼70% of patients (25 out of 37) and the remainder still unexplained. Homozygosity for the founder LTBP2 p.R299X mutation resulted in a more severe clinical phenotype and poorer outcome despite a markedly higher number of surgical interventions. The genetically homogeneous group of p.R299X homozygotes showed variable phenotypes (presumably also underlying pathogenetic mechanisms), wherein PCG proper with primary dysgenesis of the trabecular meshwork, and Marfan syndrome-like zonular disease with ectopia lentis and later onset secondary glaucoma are two extremes. The spectrum manifestations may occur in different combinations and have a different evolution even within the same sibship or a single patient. Preliminary observations on compounds with mutations in both CYP1B1-LTBP2 suggest that the observed combinations are of no clinical significance and digenic inheritance is unlikely. We provide a population genetics perspective to explain the allelic heterogeneity, comparing the history and geographic distribution of the two major founder mutations – p.R299X/LTBP2 and p.E387K/CYP1B1.

Genome-wide association studies have reported the role of the interleukin (IL) 2-IL21 chromosomal region at 4q27 in several autoimmune conditions. Mice deficient in IL-2 develop a disease with clinical and histological similarity to ulcerative colitis (UC) in humans. Modest evidence of linkage with UC was tentatively proposed for the IL2 gene more than a decade ago. Therefore, we decide to investigate the association of polymorphisms in the IL-2 axis (IL2, IL2RA, and IL2RB genes) with inflammatory bowel diseases (IBDs).Seven hundred and twenty-eight white Spanish unrelated IBD patients (356 Crohn's disease (CD) and 372 UC) and 549 ethnically matched controls were included in a case-control study. In addition, a Spanish replication cohort with 562 CD and 430 UC patients and 1,310 controls were analyzed. Eight single-nucleotide polymorphisms previously associated with different autoimmune diseases were analyzed using TaqMan chemistry.The IL2-rs6822844 polymorphism modified CD predisposition (P=0.002; odds ratio, OR (95% confidence interval, CI)=0.61 (0.44-0.84)); this was replicated in the other Spanish cohort, resulting in a strong protective effect of the minor allele in the merged samples (P=0.0002; OR (95% CI)=0.70 (0.58-0.85)). A similar effect of rs6822844 was detected for UC. Another marker, rs11938795, also showed evidence of an association with CD (P=0.006; OR (95% CI)=0.73 (0.58-0.92)).Polymorphisms within the IL2-IL21 linkage disequilibrium (LD) block show a novel association with IBD, this is concordant with suggestive previous results of whole genome analyses in CD and type 1 diabetes. Our data agree with the effect previously observed for other conditions and delineate a shared underlying mechanism.

Independent genome-wide association studies highlighted the function of CLEC16A/KIAA0350 polymorphisms modifying the risk to either multiple sclerosis (rs6498169) or type 1 diabetes (rs2903692). This C-type lectin gene maps to a linkage disequilibrium block at 16p13 and a functional role of this gene could be envisaged for other immune-related conditions, such as inflammatory bowel disease (IBD). The present study, aimed at investigating the association of those two polymorphisms with IBD, included 720 IBD patients and 550 ethnically matched healthy controls. The effect of rs2903692 previously described in diabetes was observed specifically for Crohn's disease (CD) patients lacking the main susceptibility factor described to date, that is, three polymorphisms within another pattern recognition gene, NOD2/CARD15 (NOD2(-) vs NOD2(+) CD patients, G vs A: P=0.008; OR (95% CI)=1.54 (1.10-2.15); NOD2(-) CD patients vs controls: P=0.008; OR (95% CI)=1.37 (1.08-1.73)). Replication of these findings was performed in independent Spanish cohorts of 544 IBD patients and 340 controls and the combined data yielded significant differences (405 NOD2(-) vs 204 NOD2(+) CD patients, G vs A: P=0.0012; OR(M-H) (95% CI)=1.49 (1.17-1.90); NOD2(-) CD patients vs controls: P=0.0007; OR(M-H) (95% CI)=1.35 (1.13-1.60)). The pooled analysis of the ulcerative colitis patients vs controls also yielded a significant risk (P=0.0005; OR (95% CI)=1.52 (1.19-1.93)). These data would suggest that microbial recognition through different pathways seems to converge in the development of these polygenic bowel diseases.

The longer alleles (CA)17-20 of the promoter region of PARP-1 gene may confer some protection against bilateral Meniere's disease (BMD).To analyze microsatellite (CA)(n) polymorphisms in the promoter region of PARP-1 gene and seek out risk and protective variants for BMD.Eighty patients from two ethnically defined groups with definite BMD, according to the diagnostic scale of the American Academy of Otolaryngology Head and Neck Surgery, were compared with a group of 371 normal controls from the same origin in a prospective multicenter study. We developed a specific amplification protocol to determine the PARP1-promotor CA microsatellite polymorphisms.We found that the longer alleles (CA)17-20 had a very low frequency in BMD (2/160, 1.3%, OR=7.33 (1.77-30.37, 95% CI), corrected p=0.012), suggesting that it may confer some protection against BMD.

Crohn's Disease is one of the two major forms of the Inflammatory Bowel Diseases and, although the etiology is not completely understood, the confluence of environmental and genetic factors has been demonstrated. The aim of this study was to determine the distribution of TLR4 variants in a Spanish cohort of Crohn's Disease patients and their relation with phenotype and common NOD2 variants. A total of 371 Crohn's Disease (CD) patients and 636 healthy controls (HC) were included. Single Nucleotide Polimorphisms (SNPs) in TLR4 (D299G and T399I) and NOD2 (R702W and G908R) detection was performed by a Taqman(®) Allelic Discrimination Assay. 1007insC NOD2 variant was analyzed using a PCR combined with fluorescent technology and the different alleles were determined depending on the PCR products size. D299G and T399I were related to CD only in patients carrying NOD2 variants (NOD2+/TLR4+ haplotype) (p=0.036; OR=1.924), increasing the risk to develop CD when 1007insC and TLR4 variants were both present (OR=4.886). We also described a strong association between mutant NOD2 and CD risk (p

Summary Ulcerative colitis (UC) is one of the two major forms of inflammatory bowel disease, the aetiology of which remains unknown. Several studies have demonstrated the genetic basis of disease, identifying more than 130 susceptibility loci. The major histocompatibility complex class I chain-related gene A (MICA) is a useful candidate to be involved in UC pathogenesis, because it could be important in recognizing the integrity of the epithelial cell and its response to stress. The aim of this study was to analyse the relationship between polymorphisms in the transmembrane domain of MICA and susceptibility to develop UC. A total of 340 patients with UC and 636 healthy controls were genotyped for MICA transmembrane polymorphism using a polymerase chain reaction (PCR) combined with fluorescent technology. Different MICA alleles were determined depending on the PCR product size. The allele MICA*A4 was less frequent in patients than in controls (P = 0·003; OR = 0·643), and this protective role is higher when it forms haplotype with B*27 (P = 0·002; OR = 0·294). The haplotype HLA-B*52/MICA*A6 was also associated with UC [P = 0·001; odds ratio (OR) = 2·914]. No other alleles, genotypes or haplotypes were related with UC risk. Moreover, MICA*A5.1 is associated independently with abscesses (P = 0·002; OR = 3·096) and its frequency is lower in patients diagnosed between ages 17 and 40 years (P = 0·007; OR = 0·633), meaning an extreme age on onset. No association with location, extra-intestinal manifestations or need for surgery was found.